Single Molecule Fret Studies Of Protein Conformational Landscapes: 3 Prototypic Examples For The Relation Between Conformational Dynamics And Function

Since the major structural biology method, X-ray crystallography, is limited to resolving fixed homogeneous conformations, techniques like single molecule FRET are becoming more important to determine dynamic behaviour of proteins. In comparing three different single molecule FRET studies of prototypic proteins we show that their structural aspects and dynamic behaviour are closely related to function.Initiation factor 3 (IF3) plays an important role in protein synthesis since it is an activating ligand for the correct assembly of a ribosome. It could be shown, that free IF3 is a highly flexible molecule. However this flexibility is lost completely after binding to the 30s subunit of the ribosome where it functions as a structural stabiliser, supporting the conformer selection model for ligand binding.Human guanylate binding protein 1 (hGBP1) is a member of the dynamin superfamily which is known for mediating membrane fusion in endocytosis. We show fast dynamic processes in solution between two distinct conformations. After binding its substrate GTP, hGBP1 becomes more rigid, working as a conformational switch where the GTP-bound conformation allows dimerisation leading to a functional dimer.DNA polymerases, such as the Klenow fragment of DNA polymerase I, are highly processive enzymes synthesizing double stranded DNA with a rate constant, kpol, of ∼40 s-1. We could show fast dynamics between different conformations of the enzyme in the absence of substrate. After binding of DNA and the “correct” dNTP, a closure of the protein is expected but however was not seen, rather the equilibria is shifted towards a closed conformation. This flexibility reflects the balance between specificity and velocity for this polymerase.